Single-molecule imaging is a promising method for direct quantification of the dynamics and distribution of biomolecules in living cells. Although numerous methods have been developed to gain
biological insights into molecular behavior, the high diversity of microscopes and single-molecule
dynamics can result in incomplete reproducibility of analyses. Here, we present Slitflow, an open-source framework for a single-molecule analysis workflow that includes image processing, dynamics
analysis, and figure creation. We demonstrated the integrity and flexibility of the workflow using (1)
a cherry-picked tracking method combining popular tools and (2) various state-of-the-art analyses in
a single pipeline. The software accommodates a large variety of data and methods, paving the way for
integrative analyses.
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